An analysis of regulation of phospholipid synthesis will be conducted in the unicellular eukaryotic organism, Saccharomyces cerevisiae. The studies will involve isolation and characterization of mutants with defects in the synthesis and/or regulation of phospholipids. The mutants will be employed in regulatory studies and they will be used to facilitate the cloning of structural genes encoding membrane associated enzymes of phospholipid biosynthesis. The cloned genes will be employed in studies of the regulation of phospholipid synthesis in response to exogenous precursors, such as inositol and choline. The cloned genes will also be used to facilitate studies of the assembly of these proteins into their specific locations in the cellular membranes. Little is known about the mechanisms by which eukaryotic cells control the synthesis of membrane lipids in coordination with ongoing membrane biogenesis. Yet phospholipids, particularly inositol containing phospholipids have been implicated in complex signalling processes which play a role in controlling cell growth and proliferation in higher eukaryotes. Yeast cells synthesize a typically eukaryotic mixture of phospholipids, using pathways which are similar to those in higher eukaryotes. The organism is genetically tractable, and can be manipulated using a powerful molecular genetics. Therefore, S. cerevisiae presents an unusual opportunity to develop a comprehensive understanding of the regulatory mechanisms controlling phospholipid synthesis during membrane biogenesis and cell growth in a simple eukaryote.